The present invention relates to avenanthramides, and more specifically, to a method for increasing concentration of avenanthramides in oats.
Oats are unique amongst the cereals in their capacity to synthesize and store several groups of bioactive phytochemicals including avenanthramides, avenacosylates, triterpene saponins (avenacosides) and steroid saponins (avenacins).
Avenanthramides are a group of about 30 different N-aroylanthranilic acid alkaloids of the general formula depicted in FIG. 1.
Avenanthramides are antioxidants and have been shown to be bio-available in humans and other animals, to have cardiovascular health effect in vivo.
Using purified avenanthramide mixtures, it has been shown that avenanthramides are bio-available in hamsters and humans, and act synergistically with vitamin C to enhance LDL-cholesterol resistance oxidation. In another study, the pharmacokinetics of avenanthramide uptake in healthy adults was determined. Using a randomized, placebo controlled, three way crossover trial with purified avenanthramides, at two oral dose levels, approximately 60 and 120 mg, avenanthramides were bio-available and enhanced the oxidative defense system of humans, in a dose-dependent manner, as measured by plasma glutathione levels. It can be concluded that for the observed bio-activity, threshold levels of avenanthramides, approximately 30 to 60 mg from a dietary source delivery system such as a 50 g serving of oat bran, would require an oat product with about 600 to 1,200 parts per million (ppm) total avenanthramides. This is a substantially higher concentration than those currently recorded for existing oat varieties or existing oat products.
It is well known that oatmeal, usually formulated into colloidal suspensions have been used topically for hundreds of years for skin conditions such as eczema, poison ivy, insect bites, sunburn, and shingles, where inflammation is known to be the main culprit. Avenanthramides at about 10 ppm now form the core active ingredient in a number of products for personal and pet care markets worldwide due to their topical anti-irritant, anti-itch and anti-inflammatory activities.
Avenanthramides also have shown anti-cancer preventative properties in vitro. Synthetic avenanthramide C (FIG. 1; n=1, R1═OH, R2═H, R3═OH,) at about 40 ppm has recently been shown in vitro to inhibit the proliferation of smooth muscle cells and nitric oxide production in both rat and human embryonic aorta vessel wall cultures. At concentrations of 4 to 20 ppm in this in vitro system, an avenanthramide mixture purified from VAO-6 bran significantly reduced the “stickiness” of these cells towards oxidized LDL-cholesterol and inhibited the formation of pro-inflammatory signaling molecules that promote the build-up of cholesterol-based arterial plaque. In addition, it has recently been found that purified oat avenanthramides, avenanthramide C in particular have anti-inflammatory and anti-proliferative activity against colonic cancer cell lines when treated at about the 20 to 50 ppm level, but had no effect on normal cell lines.
Avenanthramides are also potent antioxidants in vitro. It has been shown that avenanthramides are inhibitors of certain pathogenic stages of atherosclerosis (cardiovascular disease), the leading cause of morbidity and mortality in Western society. Using human aorta epithelial cell cultures, both individual avenanthramides and purified oat avenanthramide mixtures extracted from oat bran showed anti-atherogenic and anti-inflammatory bioactivity. More recently, the mechanism of the anti-inflammatory action of avenanthramides in human aortic monolayer cell cultures has been attributed to their inhibition of nuclear factor κB activation by inhibiting the phosphorylation of IκB kinase and Iκκ proteins, key factors in the initiation, progression and complication of atherogenesis.
Avenanthramides are present in current oat varieties at concentrations too low to elicit these benefits. From clinical evidence involving human subjects, effective minimum levels of single doses would have to be in the range of 1000 to 3000 ppm antioxidant.
However, North American and Scandinavian covered oat varieties generally contain from about 4 ppm to about 150 ppm total avenanthramides and these levels can vary widely depending on genotype, environment, crop year and location. Regardless of genotype/environment interactions, the avenanthramide content of outer layers of the grain is always higher than that of the starchy endosperm indicating avenanthramides are localized primarily in the bran fraction. Prior art cultivation efforts show levels can be increased up to about 130% of the starting levels in dry kernels.
The levels of avenanthramides in whole kernels can also be increased through physiological and/or mechanical processes. Malting is a process of soaking and germinating cereal grains to change the composition of the grain for a variety of end purposes, and has been practiced for millennia. For example, the malting process leads to the breakdown of complex carbohydrates, lipids and protein, rendering these reserve sources of sugars, fatty acids and amino acids more nutritionally available both to the grain itself, for further development of the embryonic plant, and to those organisms consuming it.
For example, it has been shown that the total amount of avenanthramides increased 150% during a 48 hours germination period. It has also been shown that total avenanthramides increased in whole oat kernels from about 90 ppm to about 110 ppm, a 27% increase, during 10 hours of steeping in tap water at 20° C. The increase in total avenanthramide levels was time and temperature dependent. With increased temperature of steeping, levels increased by as much as 50% to about 75 ppm during 10 hours of steeping. Increasing the temperature above 20° C., for example at 40° C., or prolonging steeping, for example, for 48 hours, did not result in further increases in avenanthramides. Simply initiating water imbibitions in dry kernels results in avenanthramide increases although the levels increase only marginally, and did not result in accumulation of substantial quantities.
It is apparent from the existing studies that current technologies for increasing avenanthramides through malting will not produce a malted product with sufficiently high levels of avenanthramides to elicit the desired physiological responses outlined above. Furthermore, since the malting/germination processes which produce these increases also result in a germinated oat seedling with roots, shoots and partially-depleted malted kernels, especially with malting times in excess of 2 days, the sprouted product would have limited use in any further dry fractionation/milling application due to the presence of roots, coleoptile and other anatomical modifications associated with germinated kernels. In addition, existing technologies have been developed mainly for covered oats, resulting in a sprouted grain with hulls still attached. De-hulling prior to malting by mechanical means, such as impact or compressed air de-hulling, widely used in the industry as well as the secondary “polishing” procedures, i.e. removal of trichomes located on the outside of the oat groat, may severely compromise the integrity of the intact kernel resulting in damaged kernels which is undesirable for malting. Furthermore, de-hulling and polishing of the malted product is complicated by changes in the softness and density of the malted material.
The germinated oat sprouts consisting of roots, shoots and partially-depleted malted kernels are also difficult to mill into either traditional oat products such as rolled oat flakes, oat bran or oat flour, suitable for incorporation into directly-consumable foods or for incorporation into baked foods as an ingredient.
In addition, given the strong genotype/environment interactions already observed, the possibility of producing a high avenanthramide oat through classical breeding techniques represents a long-term solution which might take many years to achieve.
Therefore, there is a need for a substantial increase of the concentration of avenanthramides in oat kernels so that the potential beneficial effects of avenanthramides such as in ameliorating cardiovascular health can be realized.